Abstract Although rare genetic disorders directly impact relatively small segments of the population, they are caused by mutations in genes with such critical importance that perturbed function is rarely tolerated, and therefore offer unique insight into fundamental cellular mechanisms. One such disease, fibrodysplasia ossificans progressiva (FOP), is caused by misregulated control of cell fate decisions that leads to congenital skeletal malformations and disabling extra-skeletal (heterotopic) endochondral ossification (HO) that often forms in response to tissue injury. Notably, this de novo bone formation is associated with an impaired muscle repair response. We identified that all familial and sporadic cases with classic features of FOP carry the same heterozygous mutation in ACVR1/ALK2 (R206H; c.617G>A), a cell surface receptor that mediates signal transduction of bone morphogenetic proteins (BMPs). Our data showed that the ACVR1 R206H mutant receptors mildly activate the BMP signaling pathway in the presence or absence of BMP ligands. This proposal seeks to identify how the resulting gain of function in ACVR1/BMP signaling diverts the program of muscle repair from one that normally culminates in restoration of muscle tissue to one in which muscle injury leads to differentiation of endogenous mesenchymal progenitor cells (MSCs) to chondrocytes and osteoblasts and the formation of heterotopic bone tissue. Previous studies confirmed cell autonomous effects of the mutation on MSC differentiation, however, while the mutation enhances MSC chondro/osteogenesis, we have also established that mutant cells do not spontaneously differentiate, but require additional signals. Since commitment and differentiation of tissue-resident progenitor cells is regulated by signals from the tissue microenvironment, and the tissue microenvironment is itself defined by matrix production by these differentiating cells, this proposal focuses on how enhanced BMP pathway signaling in FOP changes cellular interpretation and fabrication of the biomechanical environment during muscle repair. Based on our preliminary data showing altered physical (mechanical) properties of mutant skeletal muscle tissue following injury, this proposal will first investigate and identify the mechanisms (cellularity, matrix, and stiffness) through which ACVR1 R206H mutant tissue alters the connective tissue microenvironment during the early response to muscle injury (Aim 1). Next, we will examine the mechano-sensing signaling mechanisms through which chondro/osseous mesenchymal (non-myogenic) progenitor cells (MSCs) differentially sense and interpret signals from their microenvironment (Aim 2). Finally, we will determine the effects of the mutant tissue microenvironment on endogenous myogenic muscle progenitor cells (MuSCs, Aim 3). Together, these data will identify novel mechano-regulatory mechanisms controlling cell differentiation in heterotopic ossification and muscle repair and as well as reveal new targets for therapeutic interventions to prevent genetic and non- genetic forms of HO and to engineer tissues for clinical application.